Glucose homeostasis requires the concerted efforts of numerous neuroendocrine systems. Pancreatic islets, however, are considered to be the primary "glucose sensor" in mammals. Islets contain four populations of cells which are characterized primarily by their production of insulin, glucagon, somatostatin or pancreatic polypeptide. Among these, insulin-producing .beta.-cells predominate. Insulin secretion and production are stimulated by increases in serum glucose, an event which is mandatory for subsequent glucose uptake in certain tissues. Hence, dysfunction or destruction of .beta.-cells results in elevated serum glucose levels, ultimately developing into diabetes.
Genetic linkage analysis indicates that hereditary factors strongly influence susceptibility to acquisition of the diabetic state. For example, at least 18 genetic loci have some degree of linkage to insulin-dependent diabetes mellitus (IDDM). One disease susceptibility locus, termed IDDM2, encompasses the human insulin gene and is associated with altered transcriptional regulation of insulin promoter function. Hence, disruption of the processes that regulate insulin gene expression may account in part for diabetogenesis. Consistent with this hypothesis, impaired .beta.-cell function is a very common feature of diabetes.
Non-insulin dependent diabetes mellitus (NIDDM) is thought to occur as a result of both external and complex genetic influences. Interestingly, allelic variants at the insulin locus itself have been associated with the disease. These variants appear to contain a normal insulin gene, but exhibit altered properties with regard to transcriptional regulation.
Estimates indicate that as many as 20 million Americans may suffer from Type II diabetes. The progression of the disease appears to require both environmental factors and certain as yet largely unidentified diabetes susceptibility genes, which may contribute to the peripheral insulin resistance of type II diabetics, in which tissues fail to utilize glucose appropriately in response to the insulin signal. Alternatively, genetic factors may account for the reduced glucose sensitivity of the insulin-producing pancreatic .beta.-cells in these individuals. The end result of both of these physiological states is the marked hyperglycemia which constitutes the primary hallmark of diabetes.
Transcriptional control of the insulin gene is achieved through a short region of flanking DNA that interacts with cell-specific and glucose-sensitive signalling molecules. The precise nature of this regulatory organization remains poorly understood, although it is generally acknowledged that basic helix-loop-helix (bHLH) and homeodomain-containing factors are critical components of the transcriptional machinery that governs .beta.-cell-specific expression of insulin. An islet-specific bHLH complex interacts with a proximal E-box that has been variously termed Nir, IEB1 or ICE; this element is present twice in the rat insulin I gene, but only once in the rat insulin II and human insulin genes.
Transient assays in insulin-producing cell lines suggest that E-box-binding factors synergize with .beta.-cell-specific proteins that bind a nearby AT-rich sequence termed FLAT, which bears the hallmarks of a homeodomain recognition sequence. Indeed, several characterized homeodomain proteins have been shown to bind the FLAT element, including Isl-1, lmx-1, cdx-3 and STF-1. In addition, the latter of these corresponds to the principal binding activity at an evolutionarily conserved AT-rich sequence termed the P-element. Isl-1 binds the FLAT element weakly and does not appear to be present in the FLAT-binding complexes detected with extracts from insulin-producing cells; current evidence supports a more important role for Isl-1 in neural development. The homeodomain factors imx-1 and cdx-3 have interesting transactivation properties with regard to insulin promoter function in heterologous cells, but their cellular distribution and FLAT-binding ability inside the .beta.-cell remains unclear. In addition, there is little data that directly address the function of these factors in .beta.-cell lines. Hence, there is currently no conclusive evidence that establishes any of these factors as a principal regulator of insulin gene expression.
Within the group of factors with insulin promoter-binding activity, STF-1 is perhaps the most promising candidate for a bona fide regulator of insulin promoter function. In mice, STF-1 is first detected at embryonic day 8.5 in the nuclei of primordial cells that gives rise to the pancreas, shortly prior to the earliest detected expression of insulin in this region. Throughout the ensuing development of the endocrine pancreas, STF-1 and insulin are largely coexpressed. In addition, in extracts from insulin-producing cell lines, STF-1 appears to be a component of the enidogenous DNA-binding activity at both the FLAT and P elements in the insulin promoter. STF-1 also strongly synergizes with the E-box-binding factor Pan-1, as might be expected from a FLAT-binding factor. However, DNA-binding assays indicate that other, unknown, factors from .beta.-cell extracts also make a large contribution to the detected FLAT-binding activity. It remains unclear whether FLAT-mediated insulin promoter activity requires all, or only a subset, of these detected species.
In addition to a clear role for the FLAT-binding factors in determining .beta.-cell-specific insulin gene expression, substantial evidence also implicates these factors in glucose-responsive insulin promoter function. However, there is currently no data whatsoever which evaluates the possible role of currently cloned FLAT-binding factors as mediators of the latter function.